Immunoglobulin single variable domains (as further described herein) are characterized by formation of the antigen binding site by a single variable domain, which does not require interaction with a further domain (e.g. in the form of VH/VL interaction) for antigen recognition. Immunoglobulin single variable domains against a wide range of different targets have been described (WO 04/062551, WO 05/044858, WO 06/040153, WO 06/122825, WO 07/104,529, WO 08/020,079, WO 08/074,839, WO 08/071,447, WO 08/074,840, WO 08/074,867, WO 08/077,945, WO 08/101,985, WO 08/142,164, WO 09/068,625, WO 08/142,165, WO 09/068,627) that could be candidates for drug development. Immunoglobulin single variable domains against the p19 subunit of IL-23 that block the interaction of IL-23 with its receptor have been described e.g. in WO 09/068,627. Immunoglobulin single variable domains against the F protein of human Respiratory Syncytial Virus (hRSV) that can neutralize hRSV have been described e.g. in WO 09/147,248.
Most immunoglobulin single variable domains in pre-clinical or clinical development have been administered parenterally (i.e. by intravenous or subcutaneous administration) and stable formulations for these administration methods have been described (see e.g. PCT application No. PCT/EP2010/062972 (filed 3 Sep. 2010) based on U.S. provisional application No. 61/275,816 (filed 3 Sep. 2009) and PCT application No. PCT/EP2010/062975 (filed Sep. 3, 2010) based on U.S. provisional application No. 61/284,502 filed by Ablynx N. V. on 18 Dec. 2009). These delivery methods, however, have a rather low patient acceptance and a need exists for alternative, more convenient (needle-free) modes of administration which can easily be performed by the patients themselves.
One possible alternative method is the delivery of the immunoglobulin single variable domain through the lungs. Pulmonary drug delivery can be achieved by inhalation, orally and/or nasally. Examples of pharmaceutical devices for pulmonary delivery include metered dose inhalers (MDIs), dry powder inhalers (DPIs), and nebulizers. Traditionally, nebulizers have been classified into two main types: air-jet (pneumatic) and ultrasonic devices. Recently, a third type, vibrating-mesh nebulizers has been commercialized (Newman and Gee-Turner, 2005, J. Appl. Ther. Res. 5: 29-33).
The nebulizer is a logical first choice for development of a pharmaceutical protein-based drug for pulmonary delivery as most proteins are purified and stored as aqueous concentrate. However, the stability of aqueous solutions on the shelf may not translate to stability during nebulization and the protein may be denatured by several mechanisms including drying, shear and surface effects (Charm and Wong, 1970, Biotechnol. Bioeng 12: 1103-1109; Andrews, 1991, Biochem. Soc. Trans, 272S 19). It was shown, for example, that nebulization induced the loss of enzymatic activity of lactate dehydrogenase (LDH) and resulted in aggregation, which mainly consisted of dimer formation, and degradation of recombinant human granulocyte stimulating factor (G-CSF) (Niven and Brain, 1994, Int. J. Pharm., 104: 73-85).
WO 04/041867 describes methods and compositions for the pulmonary delivery of immunoglobulin single variables domain. In WO 09/074,634, methods of direct pulmonary delivery of domain antibodies, and particular domain antibody compositions suitable for direct pulmonary delivery, are described. More specifically, compositions are described which comprise a domain antibody polypeptide and a buffer containing 2% to about 10% PEG1000 and 1.2% sucrose (w/v). These domain antibody compositions appear to have a viscosity that allows the production of sufficient droplets with the correct size for administration to a subject by direct local pulmonary delivery. None of the above documents describe or discuss the reduction of aggregate formation and/or the improvement of the stability during pulmonary delivery of immunoglobulin single variable domains. There remains a need for additional methods and compositions for the delivery of intact and functional immunoglobulin single variable domains by the pulmonary route.